Studies will continue on the structures and mechanisms of the tetrahydropterin dependent hydroxylases. The regulatory and catalytic domains of tyrosine, phenylalanine, and tryptophan hydroxylase have all been expressed separately in E. coli. The structure determination by X-ray diffraction methods of the catalytic domain of tyrosine hydroxylase in the absence and presence of substrates or substrate analogs will be completed. The effects of phosphorylation on the structure of the regulatory domain of tyrosine hydroxylase will be determined by using high field NMR methods. Chimeric proteins containing regulatory and catalytic mutants will be constructed based on these structures. The regulatory and catalytic properties of the mutant proteins will be determined.